Immunoglobulin isotype influences affinity and specificity

Despite the increasing number of multivalent antibodies, bispecific antibodies, fusion proteins, and targeted nanoparticles that have been generated and studied, the mechanism of multivalent binding to cell surface targets is not well understood. Here, we describe a conceptual and mathematical model of multivalent antibody binding to cell surface antigens. Our model predicts that properties beyond 1:1 antibody:antigen affinity to target antigens have a strong influence on multivalent binding. Predicted crucial properties include the structure and flexibility of the antibody construct, the target antigen(s) and binding epitope(s), and the density of antigens on the cell surface. For bispecific antibodies, the ratio of the expression levels of the two target antigens is predicted to be critical to target binding, particularly for the lower expressed of the antigens. Using bispecific antibodies of different valencies to cell surface antigens including MET and EGF receptor, we have experimentally validated our modeling approach and its predictions and observed several nonintuitive effects of avidity related to antigen density, target ratio, and antibody affinity. In some biological circumstances, the effect we have predicted and measured varied from the monovalent binding interaction by several orders of magnitude. Moreover, our mathematical framework affords us a mechanistic interpretation of our observations and suggests strategies to achieve the desired antibody-antigen binding goals. These mechanistic insights have implications in antibody engineering and structure/activity relationship determination in a variety of biological contexts.

“…For example, some antibodies likely bind epitopes that limit their flexibility. Other antibodies may inherently vary in their flexibility and affect the accuracy of this prediction (35).…”

Section: Discussionmentioning

confidence: 99%

A Modeling and Experimental Investigation of the Effects of Antigen Density, Binding Affinity, and Antigen Expression Ratio on Bispecific Antibody Binding to Cell Surface Targets

Rhoden

Dyas

Wroblewski

2016

“…In the complexes with different mAbs, the shifts seen in the amide resonances in NMR experiments prior to proteolysis were negligible, indicating that the amide bonds of the two labeled residues were in a similar chemical environment as they were in their Ab-free form. Future uniform 13 C, 15 N labeling of P1 may give more insight into the chemical environments and subtle differences of the mAb-binding pockets imposed by different C regions.…”

Section: Discussionmentioning

confidence: 99%

“…This tidy view of separate structural and functional domains comprising an immunoglobulin G molecule has unraveled in recent years with various observations that C regions can affect the interaction of certain V regions with their Ag (2-7). At least six independent groups have reported findings that isotype switching is associated with altered specificity despite conservation of V region sequences (7)(8)(9)(10)(11)(12)(13). However, the molecular mechanisms for these phenomena are not understood.…”

mentioning

confidence: 99%

Variable Region Identical Immunoglobulins Differing in Isotype Express Different Paratopes

Janda

Eryilmaz

Nakouzi

et al. 2012

Self Cite

Background:The mechanism by which antibody constant region alters fine specificity is unknown. Results: Different constant regions were found to change electronic and chemical properties of the antigen-binding site. Conclusion: Constant regions can affect the energy landscape of the variable region. Significance: These results are potentially critical for understanding fast, correct immune responses at the systems level and for future immunotherapy development.

“…Additionally, the arrangement of the Fab C domains relative to the V domains and to each other may increase the probability of an appropriate relative orientation of the V heavy (H) and light (L) chains, which, in turn, can shape the Ag-binding site and affect affinity. The influence of the C region on V region affinity and/or specificity has important implications for the choice of isotype in therapeutic Abs and for the creation of more effective vaccines (20). Very recently, Casadevall and co-workers (40) studied four IgG isotypes with identical V regions by small angle x-ray scattering, and they provided additional evidence that the Ig V and C domains influence each other structurally and suggested that V region structure can have significant effects on the overall Ig structure.…”

Section: Discussionmentioning

confidence: 99%

“…In contrast, numerous authors have reported that both variable (V) and constant (C) Ig regions contribute to the binding affinity and specificity of antibodies (11,(17)(18)(19)(20)(21)(22)(23). Most of these studies have focused on affinity differences between distinct Ig isotypes.…”

mentioning

confidence: 99%

Thermodynamic Analysis of the Binding of 2F5 (Fab and Immunoglobulin G Forms) to Its gp41 Epitope Reveals a Strong Influence of the Immunoglobulin Fc Region on Affinity

Crespillo

Casares

Mateo

et al. 2014

Background:Little is known regarding the thermodynamics of binding of the broadly neutralizing anti-HIV-1 mAb 2F5 to its gp41 epitope. Results: Isothermal titration calorimetry reveals strong differences between IgG and Fab. Conclusion: Residues flanking the core epitope and the immunoglobulin Fc region contribute strongly to affinity by allosteric mechanisms. Significance: The results may help to develop new therapeutics and/or vaccines against HIV and to understanding Ag-Ab recognition.